DNA alterations in ovarian adult granulosa cell tumours: A scoping review protocol

Background Identifying and describing molecular alterations in tumors has become common with the development of high-throughput sequencing. However, DNA sequencing in rare tumors, such as ovarian adult granulosa cell tumor (aGCT), often lacks statistical power due to the limited number of cases in each study. Questions regarding personalized treatment or prognostic biomarkers for recurrence or other malignancies therefore still need to be elucidated. This scoping review protocol aims to systematically map the current evidence and identify knowledge gaps regarding DNA alterations, actionable variations and prognostic biomarkers in aGCT. Methods This scoping review will be conducted based on Arksey and O’Malley’s methodological framework and later modifications by JBI Evidence Synthesis. The protocol complies with Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for scoping reviews. All original publications describing molecular alterations of aGCT will be included. The search will be performed in May 2024 in the following databases: MEDLINE (Ovid), Embase (Ovid), Web of Science Core Collection and Google Scholar (100-top ranked). Discussion This scoping review will identify knowledge and gaps in the current understanding of the molecular landscape of aGCT, clinical trials on actionable variations and priorities for future research. As aGCT are rare, a possible limitation will be the small sample sizes and heterogenic study settings. Scoping review registration The review protocol is registered at Open Science Framework under https://doi.org/10.17605/OSF.IO/PX4MF.


Background
Ovarian adult granulosa cell tumor (aGCT) is a rare tumor arising from the ovarian stroma and accounts for approximately 2-5% of ovarian malignancies [1,2].Most aGCTs are diagnosed at an early stage and treated curatively with surgical resection.Unfortunately, approximately 10-20% relapse with advanced tumor spread, sometimes many years after initial diagnosis [3,4].In addition, aGCT patients have a lifetime increased risk for other cancers, primarily estrogen-sensitive cancers (i.e.breast and endometrial cancer) [5,6].The molecular landscape of breast and endometrial cancers has been extensively researched, yet no common driver for mutations in breast, endometrial and aGCT has been identified [7].To our knowledge, there is only one case report examining DNA variants found in women with aGCT and concurrent endometrial cancer [8].
Optimal management of aGCT presents significant challenges.Apart from the tumor stage, no prognostic biomarkers are used for prediction of potential recurrences [9,10].There is a lack of evidence-based treatment options for recurrent aGCT, except further surgery, and limited experience with targeted therapy [11][12][13][14][15][16].Finally, there is sparse information published about determining a woman's risk of other cancer diagnoses after an aGCT is identified.
Due to the tumour's rarity, molecular studies on aGCT are naturally limited to a small number of cases or extensive cross-sectional studies.These study designs cannot answer critical questions about the linkage between the genomic landscape and prognostic or actionable targets.To our knowledge, none of the existing molecular variants in aGCT are targetable for personalized treatment.Although several reviews on aGCT exist [31][32][33][34][35][36][37][38], none have systematically mapped the current knowledge of DNA variants in aGCT.
This scoping review aims to systematically describe the DNA variations in aGCT and reference these variants with well-established genetic variant databases.By referencing variations with genetic variant databases, we can report on the variant's effect on disease development and the potential for targeted therapies.

Methods and design
This review will be conducted in accordance with the methodology outlined by Arksey and O'Malley [39], and amendments proposed by the Joanna Briggs Institute (JBI) [40].The review protocol is registered at Open Science Framework under this link: https://doi.org/10.17605/OSF.IO/PX4MF.The scoping review will be developed in five stages [39]: • Stage 1: Defining the research question.
• Stage 2: Identification of relevant studies.
• Stage 3: Selection of the studies.
• Stage 4: Organisation and tabulation of the data.
• Stage 5: Summarisation, compilation and documentation of the results.This protocol was developed in accordance with the guidelines from the PRISMA Extension for Scoping reviews (PRISMA-ScR) checklist [41] (S1 and S2 Checklists).

Stage 1: Identification of the research question
Inspiration for this review was elicited from a recently published scoping review on the molecular alterations in peritoneal mesothelioma (PeM) [42].This scoping review identified common mutations in rare cancers and used MyCancerGenome.orgas the reference for actionable targets or clinical trials [43].MyCancerGenome.org,OnkoKB, ClinVar, COSMIC, Clinical-Trial.govare databases that store information about oncogenic mutations, targetable mutations and ongoing clinical trials regarding targeted therapy [43][44][45][46].These databases will also be used as a reference for any molecular alterations identified in this scoping review.This scoping review aims: • To explore the DNA alterations associated with aGCT and give an overview of potential treatment possibilities • To investigate if reported DNA variations predict risk of recurrence, aggressive disease or a risk of developing other primary malignant tumors • To reference molecular alterations found in aGCT with MyCancerGenome.org,OnkoKB, ClinVar, COSMIC, ClinicalTrial.gov (May 2024) • To identify knowledge gaps between DNA alterations of aGCT and prognosis potential

Stage 2: Identification of the relevant literature
The authors have developed a set of inclusion criteria based on the 'Population-Concept-Context (PCC)' framework proposed by JBI [47].
Search strategy.The search strategy will consist of two search terms (granulosa cell tumour AND molecular alterations).Each element will consist of database-specific subject headings (MeSH), and free text words with relevant truncation and proximity operators.The search terms were inspired by similar search strings published in reviews from the Cochrane Library [16,48].An example of the search strategy for Medline (OVID) is presented in Box 1.
The search strategy will be translated for Embase (Ovid), Web of Science and Google Scholar (100-top ranked) and reviewed with an information specialist in health sciences.

Stage 3: Study selection
A summary of the PCC and inclusion and exclusion criteria is shown in Box 2.
Only peer-reviewed original research focusing aGCT and molecular alterations will be included.The aGCT diagnosis must have been defined and validated by pathologists prior to molecular analysis.Only articles describing high throughput sequencing techniques of somatic and germline DNA variations in women with aGCT will be considered.There will be no language or publication date restrictions, and all studies matching our criteria published up until the search date will be considered.Studies with cell lines and targeted DNA sequencing limited to only the FOXL2 variant known to be present in ~95% of aGCT will be excluded.Any reviews of molecular profiles of aGCT and the prognostic values of this information will be used to identify additional primary studies by applying forward and backwards citation searching.
Results will be imported into covidence [49] for screening, and any duplicates will be removed.Two reviewers will screen titles and abstracts, applying inclusion and exclusion criteria (Box 1).Any full-text articles excluded after screening will include the reasoning behind exclusion.A PRISMA flowchart will summarise the search, screening and identification Box 1. Search syntax.

Stage 4: Charting the data
The authors will use a chart or table (based on the JBI template source of evidence details, characteristics and results extraction instrument) to extract data blinded to each other [47].Any somatic and germline DNA alterations, known as either pathogenic, likely pathogenic or of unknown significance according to OncoKB Cancer Gene List will be recorded [44].Data to calculate frequencies will be plotted in STATA Release 17.0 (StataCorp, College Station, TX, USA).Relevant information will be presented in pre-planned tables (see Tables 1, 2 and 3 and Fig 1).Tables 2 and 3 will be generated by referencing any DNA pathogenic and likely pathogenic alterations with established databases (MyCancerGenome.org,OnkoKB, ClinVar, COSMIC, ClinicalTrial.gov) to identify the associated risks of other neoplasms and the possibility of targeted therapies.These proposed tables are preliminary and may be amended as the scoping review progresses.

Stage 5: Collating, summarizing and reporting results
The study's outcome will be published as a scoping review article containing texts, flow charts and tables.The flow chart will present the search strategy and study selection results.The identification and findings of the selected studies will be elaborated on in the article's discussion section.Moreover, the research questions will be addressed based on these main findings.Finally, limitations, knowledge gaps and areas requiring further research will be highlighted.

Patient and public involvement
No patient involvement.

Discussion
The proposed scoping review article will systematically map the current knowledge about the DNA alterations in aGCT and the associated clinical impacts of these alterations.Therefore, the focus will be on pathogenic and likely pathogenic variations, as these may impact treatment options and disease development.Only one common missense mutation in FOXL2 (c.402C>G; p.C134W) has been found in ~95% of aGCT [22][23][24].Apart from this mutation, reported somatic variants are heterogenous, and little is known about the clinical impact of these variants in aGCT [18,20].Collating the results of studies will identify common and recurrent molecular DNA variants describing sub-groups of patients with different risk profiles.This review allows for the comparison of results across publications to identify new common variants.This comparison will assist in identifying knowledge gaps and priorities for clinical trials on actionable variations in aGCT.In addition, any DNA alterations eligible for personalized treatment, including immunotherapy, might be identified by referencing the alteration with well-established databases (MyCancerGenome.org,OnkoKB, ClinVar, COS-MIC, ClinicalTrial.gov).
This scoping review methodology was previously used to map the molecular landscape in other rare tumors [42,50,51].Limitations of this scoping-review approach include the broad scope and heterogeneity of studies reviewed.Studies may lack complete mutational data or clinical outcomes such as morbidity and mortality, hampering interpretation.Although NGS Box 2. Summary of PCC.Any disagreements will be solved by discussion, or if necessary, an experienced third author will make the final decision.Screening and study selection will be performed in March 2024.

Table 2 . Pathogenic variants in genes associated with an increased risk of other primary cancers.
granulosa cell tumor; P, pathogenic; LP, likely pathogenic; VUS, variance of unknown significance.https://doi.org/10.1371/journal.pone.0303989.t002encompasses targeted gene-panel sequencing, whole exome and whole genome sequencing, each technique has strengths and limitations.Any conclusions from studies based on different molecular techniques should be cautious in interpretation.